Our studies to date have demonstrated that inhibiting RAS reduces the radiation survival of tumor cells in which RAS is activated by mutation or epidermal growth factor receptor (EGFR) signaling. We have used a pharmacological approach to inhibit RAS post-translational processing. This processing is required for RAS membrane binding and activity. We have shown that farnesyltransferase inhibitor (FTI) treatment radiosensitizes tumor cells with H-ras mutations without altering the radiation sensitivity of normal cells. While the effect was significant in tissue culture, animal experiments led us to examine whether FTI treatment had additional effects in vivo. Subsequent studies revealed that FTI treatment led to increased oxygenation in tumors xenografts expressing oncogenic H-ras. We propose to further explore the ability of FTIs to alter the tumor microenvironment by determining whether mutant RAS is the target for this action of FTIs. We will investigate the mechanism through which FTIs alter the oxygenation of tumors and whether it involves changes in tumor vasculature. We have also shown, using a genetic approach, that oncogenic H-, K-, and N-RAS all contribute to radiation resistance. Signaling from the three RAS isoforms to down-stream effectors has been reported to differ. This raises the question of which pathways downstream of RAS contribute to the observed radiation resistance. We propose to explore the contribution of each RAS isoform to intrinsic radiation resistance and to investigate downstream pathways using both pharmacologic inhibitors as well as novel genetic reagents. Small interfering RNA (siRNA) will be used to specifically target expression of the different RAS isoforms as well as to specifically target oncogenic RAS expression. The results of these studies could be valuable in developing new molecular targets to improve the efficacy of radiotherapy. They may also provide a rationale for the application of FTIs in the treatment of tumors with RAS activation that is due to EGFR signaling. [unreadable] [unreadable]